Photodynamic therapy for breast cancer in a BALB/c mouse model · Objective: Photodynamic therapy...
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INTRODUCTION There were records that ancient civilizations in Egypt, India, China, and Greek employed sun exposure to treat skin dis- eases, rickets and even psychosis. However, it has not been employed widely in medicine until the eighteen century [1]. In 1903, The group of Von Tappeiner found favorable result after photodynamic therapy (PDT) using several dyes: eosin, fluo- rescein, sodium dichloroanthracene disulfonate and ‘Grubler’ s Magdalene red’ with light on skin diseases, such as lupus of the skin and condylomata of the female genitalia [2]. Von Tappeiner and Jodlbauer [3,4] proved the necessity of oxygen in this phenomenon in 1904 and in 1907, they explained this phenomenon with oxygen-dependent photosensitization and adopted a term “photodynamic therapy”. Although these showed good results, no further reports were forthcoming because of the emergence of ionizing radiation in cancer therapy. Diamond et al. [5] reported in 1972 that hematopor- phyrin activated by white light may lead to regression of an experimental glioma in rats. Finally, Dougherty et al. [6] initi- ated clinical trials of PDT in 25 patients with cutaneous and subcutaneous tumors and confirmed the effectiveness of PDT. Original Article Photodynamic therapy for breast cancer in a BALB/c mouse model Tae-Gyu Ahn 1 , Byoung-Rai Lee 2 , Eun-Young Choi 1 , Dong Won Kim 3 , Sei-Jun Han 1 Departments of 1 Obstetrics and Gynecology and 2 Biochemistry, Chosun University School of Medicine, Gwangju; 3 Department of Obstetrics and Gynecology, Mirae and Heemang Women’s Hospital, Gwangju, Korea Received Nov 11, 2011, Revised Dec 12, 2011, Accepted Dec 20, 2011 This abstract was adopted as a poster presentation in 2011 Asian Society of Gynecologic Oncology 2 nd Biennial Meeting. Correspondence to Sei-Jun Han Department of Obstetrics and Gynecology, Chosun University School of Medicine, 309 Pilmun-daero, Dong-gu, Gwangju 501-759, Korea. Tel: 82-62- 220-3085, Fax: 82-62-232-2310, E-mail: [email protected] pISSN 2005-0380 eISSN 2005-0399 Copyright © 2012. Asian Society of Gynecologic Oncology, Korean Society of Gynecologic Oncology This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited. www.ejgo.org Objective: Photodynamic therapy (PDT) has been used for superficial neoplasms and its usage has been recently extended to deeper lesions. The purpose of this study was to observe whether or not PDT can cure breast cancer in the solid tumor model, and to define the critical point of laser amount for killing the cancer cells. Methods: Twenty four BALB/c mouse models with subcutaneous EMT6 mammary carcinomas were prepared. Mice were divided into eight groups depending on the amount of illumination, and the tumor size was between 8 mm and 10 mm. We began by peritoneal infiltration with a photosensitizer 48 hours prior to applying the laser light, and then we applied a non- thermal laser light. The energy was from 350 J/cm 2 to 30 J/cm 2 to the cancer. Results: Regardless of the tumor size from 8 mm to 10 mm, all mice apparently showed positive results via PDT. We also did not find any recurrence over 90 J/cm 2 . In all models, the color of the breast cancer lesions began to vary to dark on 2 days post PDT and the tumor regression began simultaneously. Also, we confirmed the complete regression of the breast cancer 21 days after PDT. Conclusion: We confirmed that PDT may treat breast cancers that are sized less 10 mm in mouse models. The moderate energy to destruct the breast cancer cells may be 90 J/cm 2 . Therefore, we can expect that PDT may be utilized to treat breast cancer, but we need more experience, skills and processing for clinical trials. Keywords: Breast neoplasms, Inbred BALB C, Mice, Photodynamic therapy J Gynecol Oncol Vol. 23, No. 2:115-119 http://dx.doi.org/10.3802/jgo.2012.23.2.115
Transcript of Photodynamic therapy for breast cancer in a BALB/c mouse model · Objective: Photodynamic therapy...
INTRODUCTION
TherewererecordsthatancientcivilizationsinEgypt, India,China,andGreekemployedsunexposuretotreatskindis-eases,ricketsandevenpsychosis.However, ithasnotbeenemployedwidelyinmedicineuntiltheeighteencentury[1].In1903,ThegroupofVonTappeinerfoundfavorableresultafter
photodynamictherapy(PDT)usingseveraldyes:eosin,fluo-rescein,sodiumdichloroanthracenedisulfonateand‘Grubler’sMagdalenered’with lightonskindiseases,suchas lupusoftheskinandcondylomataofthefemalegenitalia[2].VonTappeinerandJodlbauer[3,4]provedthenecessityofoxygeninthisphenomenonin1904andin1907,theyexplainedthisphenomenonwithoxygen-dependentphotosensitizationandadoptedaterm“photodynamictherapy”.Althoughtheseshowedgoodresults,nofurther reportswere forthcomingbecauseof theemergenceof ionizing radiation incancertherapy.Diamondetal.[5]reportedin1972thathematopor-phyrinactivatedbywhitelightmayleadtoregressionofanexperimentalgliomainrats.Finally,Doughertyetal. [6] initi-atedclinicaltrialsofPDTin25patientswithcutaneousandsubcutaneoustumorsandconfirmedtheeffectivenessofPDT.
Original Article
Photodynamic therapy for breast cancer in a BALB/c mouse modelTae-Gyu Ahn1, Byoung-Rai Lee2, Eun-Young Choi1, Dong Won Kim3, Sei-Jun Han1
Departments of 1Obstetrics and Gynecology and 2Biochemistry, Chosun University School of Medicine, Gwangju; 3Department of Obstetrics and Gynecology, Mirae and Heemang Women’s Hospital, Gwangju, Korea
Received Nov 11, 2011, Revised Dec 12, 2011, Accepted Dec 20, 2011
This abstract was adopted as a poster presentation in 2011 Asian Society of Gynecologic Oncology 2nd Biennial Meeting.
Correspondence to Sei-Jun HanDepartment of Obstetrics and Gynecology, Chosun University School of Medicine, 309 Pilmun-daero, Dong-gu, Gwangju 501-759, Korea. Tel: 82-62-220-3085, Fax: 82-62-232-2310, E-mail: [email protected]
pISSN 2005-0380 eISSN 2005-0399
Copyright © 2012. Asian Society of Gynecologic Oncology, Korean Society of Gynecologic Oncology
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
www.ejgo.org
Objective:Photodynamictherapy(PDT)hasbeenusedforsuperficialneoplasmsanditsusagehasbeenrecentlyextendedtodeeperlesions.ThepurposeofthisstudywastoobservewhetherornotPDTcancurebreastcancerinthesolidtumormodel,andtodefinethecriticalpointoflaseramountforkillingthecancercells.Methods: Twenty fourBALB/cmousemodelswithsubcutaneousEMT6mammarycarcinomaswereprepared.Miceweredividedintoeightgroupsdependingontheamountofillumination,andthetumorsizewasbetween8mmand10mm.Webeganbyperitoneal infiltrationwithaphotosensitizer48hourspriortoapplyingthelaser light,andthenweappliedanon-thermallaserlight.Theenergywasfrom350J/cm2to30J/cm2tothecancer.Results: Regardlessofthetumorsizefrom8mmto10mm,allmiceapparentlyshowedpositiveresultsviaPDT.Wealsodidnotfindanyrecurrenceover90J/cm2.Inallmodels,thecolorofthebreastcancerlesionsbegantovarytodarkon2dayspostPDTandthetumorregressionbegansimultaneously.Also,weconfirmedthecompleteregressionofthebreastcancer21daysafterPDT.Conclusion: WeconfirmedthatPDTmaytreatbreastcancersthataresizedless10mminmousemodels.Themoderateenergytodestructthebreastcancercellsmaybe90J/cm2.Therefore,wecanexpectthatPDTmaybeutilizedtotreatbreastcancer,butweneedmoreexperience,skillsandprocessingforclinicaltrials.
Keywords:Breastneoplasms,InbredBALBC,Mice,Photodynamictherapy
J Gynecol Oncol Vol. 23, No. 2:115-119http://dx.doi.org/10.3802/jgo.2012.23.2.115
Tae-Gyu Ahn, et al.
http://dx.doi.org/10.3802/jgo.2012.23.2.115116 www.ejgo.org
Afterthe1990s,PDThasrapidlybecomearelevantmethodbothinclinicalapplicationandbasicmechanisticunderstand-ing in treating theskinandsuperficialcancerous lesions.NowPDTwithPhotofrinhasbeenapprovedbytheUSFoodandDrugAdministration(FDA)andnumerousotherhealthagenciesthroughouttheworld forobstructiveesophagealcancerandlungcancer,bladdercancer,etc[7].RecentlyPDThasbeenclinicallyapplicableinsuperficiallesionslikeskinle-sions,cervical intraepithelial lesionsandhollowedorgans. Inourgynecologicdepartment,wehavemuchcumulativedataoftheeffectofPDToncervical intraepithelialneoplasiaandhavestartedtotreatbreastcancer.However,therearetodatenodataoftheeffectofPDTinthefieldofthebreast.SoweherebyexaminedtheresponseofbreastcancerstoPDTwithPhotofrinusingmurinemodels.
MATERIALS AND METHODS
1. Tumor modelBreastcancercells(EMT-6)wereobtainedfromtheAmeri-
canTypeCultureCollection(ATCC;Manassas,VA,USA)and
maintained inWaymouth’smediasupplementedwith15%fetalbovineserumand1%penicillin-streptomycinsolution.6-week-oldfemaleBALB/cmice(Samtako,Osan,Korea)wereanesthetizedwithketamineand1.5×105cellsweresubcuta-neouslyinjectedintothelowerback.Foreachexperiment,3micewereusedineachexperimentalgroupandtreatmentwasstartedwhentheaveragetumorsizewasapproximately250mm3.Thestudywasapprovedbythelocalanimalwelfarecommittee.
2. Protocol of PDTWebeganbyperitonealinfiltrationofphotosensitizer(Pho-
tofrin[HpD],AxcanPharmaInc.,Mont-Saint-Hilaire,Canada)with5mg/kg,48hourslater,weappliedanon-thermallaserlight(CeralasDiodeLaser632System,Biolitec,Jena,Germany)(Fig.1).Theamountofenergywasfrom350J/cm2to30J/cm2onthecancercellandthemarginoflaserwasalittlebitwiderthanthecancerlesionbecausecancercanrecureasily.Thenweevaluatedtheeffectsofthisprocedureevery3days.Thecontrolgroupwasnotneededbecauseifthebreastcancerwasnot irradiated, thegrowthofcancerwillcertainlycon-tinue.
RESULTS
JustafterPDT,graydiscolorationwasshownattheillumina-tionsite.Thecolorofilluminationsiteincludingbreastcancerbegantodarken2dayspostPDTandthecancerregressedsi-multaneously.Thetumorsizewasfrom8mmto10mm,how-ever,regardlessofthetumorsizeallcancermodelsshowedeffectivenessof thePDT.However, tissuedeformitieswerenoticedinthe240and350J/cm2group(Fig.2).Weconfirmedthecompleteremission21dayspostPDTfrom180to90J/cm2(Fig.3).From350J/cm2to90J/cm2,allmicemodelsshowedcom-
pleteremission.HoweveroneoutofthreemiceshowednoFig. 1. Photodynamic therapy illumination onto the cancer with a non-thermal laser light (Ceralas Diode Laser 932 System).
Fig. 2. Complete remission was shown in 350 and 240 J/cm2 but tissue deformities were noticed because of high energy (21 days after photodynamic therapy).
Effect of photodynamic therapy on breast cancer model
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responseinthe60J/cm2group(Fig.4)andinthe30J/cm2group,theeffectivenessofPDTdidnotshowinallmice(Fig.5).So,wedeterminedthatthemoderateenergyrequiredtotreatbreastcancersizedlessthan10mmisaround90J/cm2.Wedidnotfindanyrecurrenceinthegroupthatreceivedenergymorethan90J/cm2.So,theenergyof90J/cm2maybeminimumenergyrequiredtodestructthebreastcancercellscompletely.
DISCUSSION
PDT isa treatment thatcaneradicatepremalignantandearly-stagedcancerandcanreducethetumorsize inend-stagedcancers.TheadvantageofPDTisthatitisaminimallyinvasivemethodthatcanaffecttheselectiveareaandthereisnolimitationofrepetitionsandlittlecomplications.Itisalso
Fig. 3. Complete remission shown in 180, 150, 120, and 90 J/cm2 (21 days after photodynamic therapy).
Fig. 4. The first and third mice showed complete remission but the second one (arrow) revealed no response with 60 J/cm2 (21 days after photodynamic therapy).
Fig. 5. All mice showed no response. Cancer growth was continued with 30 J/cm2 (21 days after photodynamic therapy).
Tae-Gyu Ahn, et al.
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hasamerit inthatPDTisapplicablewhensystemicchemo-therapyandsurgeryarenotpossible[8].PDTmechanismcanbeexplainedbytwoparts:uptakeofsensitizersandtumordestructionbylaserlight.
1. PDTThefirstapprovalofPDTwasinCanadain1993forthetreat-
mentofbladdercancers.EsophagealcancerwasapprovedbytheUSFDAinDecember1995,andearlynon-smallcell lungcancer in1998followedbyBarrett'sesophagus[9,10].EventhoughPDTdataingynecologyarenotsufficient,several in-stitutesalreadyhavecollectedmuchdataandsomegyneco-logicdepartmentshavetriedtobeginPDT.Severalphotosen-sitizersthatcanbeaccumulatedinthetumortissueselectivelyhavebeen introduced,but theexactmechanismsofeachphotosensitizersaredifferent.Porphyrinderivativescanbelocalizedintumorswheninjectedintothebloodstream[11].Inthehematoporphyrinderivativeandits improvedversion,calledPhotofrin,arestillthemostwidelyusedphotosensitiz-ersinPDToftumors[12].Amongphotosensitizers,Photofrinhasbeenusedinourdepartmentforthe lasttenyearsandweassumedthatPDTwithPhotofrinmaybehaveeffectiveinbreastcancersalso. Inourdepartment,webegantooperateonbreastcancerfrom2010andwetriedtoadoptPDTinthebreast lesions.So,weconductedthisstudy.Weappliedthesameprotocolthatwasusedinpatientstothemicemodel.Thedrugisinjectedintravenouslyandafter48hours,applica-tionofirradiationtotumorswasperformedwith630nmwavelengthoflaser[13,14].
2. PDT and breast cancerPDThasbeenapplicableinthefieldsothatphysicianscan
directlyapproachthe lesion.Thebreastarea isalsoabletobeapproacheasily,andwecanadoptPDTforthebreast.Thefirst linetreatmentforbreastcancer issurgeryandcanleadtochangesinthebreastshape.Thus,ifthereisanon-invasiveprocedurethatisabletodestroycancercells,itwouldbepref-erable.EventhoughtherewerenoreportsaboutPDTforthebreastcancerasthefirstlinetherapy,severalstudiesshowedreliableeffectsofPDT in locally recurrentbreastcancerorchestwallrecurrence[15-17].Thus,wemayassumethroughthisdatathatbreastcancercellsmaybeeradicatedbyPDT.Also,PDTisapplicableduringsurgery.Aftermastectomy,PDTcanbeappliedonthebedofthepreviouslesionforprevent-ingrecurrence.TheusuallaserfibercanbeusedwhenPDTisappliedtotheskinareasuchaschestwallrecurrence.But ifwetrytotreatthelesionthatislocatedundertheskin,aspe-cificlaserfiberisneededsuchasaninterstitialtypethatcanpenetratetheskinandreachthelesion.So,researchaboutthe
specificlaserfiberismandatorytodestroylesionsundertheskin.Fromourstudy, firstwecanreconfirmthePDTeffects to
destroybreastcancercells.Next,discoveryofproperenergyisvery importantbecausetheamountofenergyshouldbedifferentforeachtissueandorgan(forexample,weapplied240J/cm2touterinecervicalcancer).Inaddition,whenphoto-chemical internalization(PCI)conceptisaddedtotraditionalPDT,theresultofcancercelldestructioncanbemaximized.ForperformingPCI, chemotherapeuticagents shouldbeaddedbeforePDT.Whenpreviouslyinfiltratedsensitizersareexposedto light,sensitizersbegintoactivateandproducefreeradicals.Finally,endocytosedmoleculesorchemothera-peuticagentsmaybereleasedandattackcancercellsdirectlybeforebeingdegradedbylysosomes[18].Thus,ifweaddthisPCIconceptwiththetraditionalPDTinournexttrial,wecanassumethattheresultswillimprovemarkedly.InourresearchwewereabletoconfirmtheeffectofPDT
onbreastcancermodelsandfoundouttheproperamountofenergyof90J/cm2inlessthan10mmsizedbreastcancers.However,weassumethatweneedmoretechnicalpracticeandclinicaldatatobeabletoapplyPDTtohumans.
CONFLICT OF INTEREST
Nopotentialconflictof interestrelevanttothisarticlewasreported.
ACKNOWLEDGMENTS
ThisresearchwassupportedbyresearchfundsfromChosunUniversity,2005.
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